Protein synthesis is a highly conserved process in all kingdoms of life that can be broken down into four distinct phases: initiation, elongation, termination and ribosome recycling. The broad goals of this research program are to use biochemical and genomic approaches to shed light on the common and distinctive molecular features of translation elongation, termination, and recycling in bacteria and eukaryotes, and their control. Here we are particularly focused on one aspect of translational control in which ribosomal stalling triggers a cellular response leading to mRNA decay, targeted proteolysis, and ribosome recycling. In particular, we focused initially on a highly conserved stalling motif, the poly-basic peptide sequence, that is of particular relevance in eukaryotic cells where alternative polyadenylation site usage commonly leads to non-stop mRNAs. We will continue to use in vitro biochemistry and in vivo ribosome profiling to look at the molecular mechanics of this biologically important and conserved process (and other related systems). More specifically, we propose (1) to use our previously established in vitro reconstituted translation system (with S. cerevisiae components) to ask a series of questions about ribosome-based mechanisms for sensing translational perturbations, (2) to use a series of reporters in yeast to screen for novel components that contribute to these mRNA surveillance pathways and (3) to use ribosome profiling approaches to define the biologically relevant in vivo targets, their molecular features, and the factors that contribute to these important pathways. We anticipate that the synergy of these approaches will be powerful in defining biologically relevant mechanism.